EP0876219B1 - Pipettage automatise de petits volumes - Google Patents

Pipettage automatise de petits volumes Download PDF

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Publication number
EP0876219B1
EP0876219B1 EP96937277A EP96937277A EP0876219B1 EP 0876219 B1 EP0876219 B1 EP 0876219B1 EP 96937277 A EP96937277 A EP 96937277A EP 96937277 A EP96937277 A EP 96937277A EP 0876219 B1 EP0876219 B1 EP 0876219B1
Authority
EP
European Patent Office
Prior art keywords
liquid
impulse generator
pipetting
diluter
tubing
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Expired - Lifetime
Application number
EP96937277A
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German (de)
English (en)
Other versions
EP0876219A1 (fr
Inventor
Anselm Sickinger
Johannes Konrad Balmer
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Tecan Schweiz AG
Original Assignee
Tecan AG
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Filing date
Publication date
Application filed by Tecan AG filed Critical Tecan AG
Publication of EP0876219A1 publication Critical patent/EP0876219A1/fr
Application granted granted Critical
Publication of EP0876219B1 publication Critical patent/EP0876219B1/fr
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Expired - Lifetime legal-status Critical Current

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Classifications

    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/02—Burettes; Pipettes
    • B01L3/0241—Drop counters; Drop formers
    • B01L3/0268—Drop counters; Drop formers using pulse dispensing or spraying, eg. inkjet type, piezo actuated ejection of droplets from capillaries
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/10—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N35/00—Automatic analysis not limited to methods or materials provided for in any single one of groups G01N1/00 - G01N33/00; Handling materials therefor
    • G01N35/10—Devices for transferring samples or any liquids to, in, or from, the analysis apparatus, e.g. suction devices, injection devices
    • G01N2035/1027—General features of the devices
    • G01N2035/1034—Transferring microquantities of liquid
    • G01N2035/1039—Micropipettes, e.g. microcapillary tubes
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T436/00—Chemistry: analytical and immunological testing
    • Y10T436/25—Chemistry: analytical and immunological testing including sample preparation
    • Y10T436/25625—Dilution
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10T—TECHNICAL SUBJECTS COVERED BY FORMER US CLASSIFICATION
    • Y10T436/00—Chemistry: analytical and immunological testing
    • Y10T436/25—Chemistry: analytical and immunological testing including sample preparation
    • Y10T436/2575—Volumetric liquid transfer

Definitions

  • This invention relates to a device for automated pipetting of small volumes of liquid according to the preamble of claim 3 and a method for automated pipetting of small volumes of liquid according to the preamble of claim 1.
  • volumes smaller than 10 ⁇ l require adjusted instruments, and most frequently, in addition new techniques of pipetting. Volumes larger than 10 ⁇ l can easily be dispensed from the air, as the drops can leave the tip by themselves if handled appropriately. For volumes smaller than 10 ⁇ l techniques have to be applied that guarantee the emission of the droplets. For this reason, the tip can touch the wall of a vessel containing the liquid to be treated or the droplet can be dispensed directly into the desired liquid. A subsequent mixing step is advantageous in order to obtain a complete transfer.
  • a third possibility, that is frequently used, is the pre-uptake of a reagent, which is finally dispensed together with the sample volume.
  • the air-displacement tip is a normal disposable tip, which is adjusted to the low volume with respect to material and geometry.
  • a piston is used inside of the tip in order to improve the accuracy.
  • diluter consists of a stepper motor driven syringe pump for precision liquid handling.
  • the diluter operates from a single 24 volt power supply and is controlled by an external computer or microprocessor. It works as slave unit automating pipetting, diluting and dispensing functions. Further particulars of this diluter are described in the Operator's manual P/N 724043C of Cavro Scientific Instruments Inc..
  • Known diluters have been developed that can aspirate and dispense volumes in the amount of nanoliters, at least in theory.
  • the accuracy (ACC) and reproducibility (CV) of the pipetting can be influenced by various parameters.
  • the most important one is the velocity of the aspiration and the dispense of the liquid. Diluters used so far are adapted to handle volumes of 10 ⁇ l with the appropriate resolution and velocity. However, the most critical point is the tearing off of the liquid from the tip, which is limited by the mechanics of the diluter. Speed of the drive can not be increased infinitely. Even the speed that is required to dispense low volumes has not yet been reached.
  • the limits are within the volume of one drop.
  • the forming, the shape and the size of a drop primarily depends on the surface tension of the liquid, which results from the chemical and physical properties of the sample of the pipetted liquid.
  • the size of a drop of water is 10 - 20 ⁇ l, which is disadvantageous for the pipetting, because this is the minimum volume, that can be torn off from the tip. This effect becomes visible while attempting to pipette volumes of 1 ⁇ l; after 12 - 15 times of dispense, the drop has reached the critical size and is emitted.
  • there do exist applications of the dispensing of tiny droplets in the amount of nanoliters for example in ink jet printers.
  • a liquid dispensing device which allows to dispense a given volume of liquid by applying a number of pressure drop pulses to the liquid.
  • US-A-5 334 353 BLATTNER a fluid delivery device is known which uses a reciprocating force mechanism acting on the tip holder of the device which is stopped in its path of travel causing the droplet on the tip to fall away.
  • EP-A-0119573 the use of an impulse generator acting via a plunger on a flexible tubing portion filled with dispensing liquid to dispense a microdroplet of liquid from the outlet orifice is known.
  • this device is usable only for dispensing and not for aspiring+dispensing, i.e.
  • the invention as claimed allows the reproducible automated pipetting of volumes of liquid smaller than 10 ⁇ l.
  • the combination of a state of the art diluter system with an impulse generator allows to shift the physical/chemical limits of the forming of drops to lower volumes in the region below 10 ⁇ l.
  • the impulse generator can be put somewhere within the liquid system and the kind of generator that is used is of minor importance. The only pre-condition is that the generator is located between the valve of the diluter and the tip, or is built into the valve, or affects the plunger drive of the diluter.
  • the device shown in Fig. 1 basically consists of a diluter 1, a pipetting needle 4, a tubing 5 connecting said diluter 1 with said needle 4, and an impulse generator 6.
  • the diluter 1 comprises a syringe 2 and a valve (3), whereby said syringe 2 comprises a piston 11 and a piston drive 9.
  • the impulse generator 6 is located between the valve 3 of said diluter 1 and said needle 4.
  • the impulse generator 6 can either be located in said valve 3 or on said piston drive 9.
  • the whole device is filled up with the appropriate system liquid, e.g. deionised water, a buffering solution or an oil.
  • a robot arm 7 moves needle 4 to the source 8, e.g. an open container with the liquid sample to be analysed. After detection of the surface of this liquid by the needle 4 and submerging it some tenth of millimetres into said liquid the predetermined amount or volume of liquid sample is aspirated from the source 8 with a definable additional "conditioning" volume of the liquid sample through the needle 4 by means of the diluter 1. After retracting needle 4 out of the source 8 the "conditioning" volume is dispensed back into the source 8 by means of the diluter 1.
  • the appropriate system liquid e.g. deionised water, a buffering solution or an oil.
  • the pipetting needle 4 has an internal diameter smaller than 0.6 mm at its free end and preferably smaller than 0.4 mm.
  • the impulse generator 6 applies a force to the liquid column in the tubing 5 to grade the remaining liquid in the pipetting needle 4.
  • the effect is - as shown in Fig. 3 - that the meniscus 18 of the liquid resides inside of the tip 4 at a level 13 that only depends of the magnitude of the impulse generated by the impulse generator 6 and the transmission of said impulse to the liquid 19 in the tubing 5.
  • the diluter 1 After moving needle 4 to the destination 10 (Fig. 1), e.g. an open container, the diluter 1 delivers the volume to be dispensed to the liquid in the tubing 5. As shown in Fig. 4 the meniscus 18 of the liquid 19 moves a distance (indicated by arrow 17) within the needle 4 to the level 14 and the impulse generator 6 again applies the impulse. The effect is - as shown in Fig. 5 - that the volume delivered by the diluter 1 (liquid column between levels 13 and 14) breaks off the tip of needle 4 in the form of drop 16 and the meniscus 18 of the liquid 19 inside the needle 4 comes to stop at the steady location at level 13 prior to the delivery of the liquid volume corresponding to drop 16. This procedure is applicable to both pipetting modes single and multi pipetting (aliquotting).
  • the steady location of meniscus 18 at level 13 depends primarily from the stroke of the impulse generator 6 and secondarily from the magnitude of the impulse generated by impulse generator 6. If constant stroke and impulse is applied to the liquid 19 the location of the meniscus 18 will always be at the steady location at level 13.
  • the impulse generator 6 can be either a mechanical impulse generator (e.g. a core magnet or a solenoid pinch valve), or an inductive impulse generator, e.g. a piezo element.
  • the impulse generator 6 can also comprise an ultrasonic source, or a heat source.
  • the magnitude of the impulses J generated by the impulse generator 6 should be in the order of 0.01 to 1.00 N ⁇ s, preferably in the order of 0.01 to 0.10 N ⁇ s.
  • the variation of the magnitude of the impulse can be achieved by altering the controlling conditions of the impulse generator 6.
  • the syringe 2 of the diluter 1 has a volume in the range of 10 - 500 ⁇ l.
  • the volume is in the range of 100 - 400 ⁇ l and typically in the range of 200 - 300 ⁇ l.
  • These indications are related to a pipetting volume in the range of 1 - 10 ⁇ l.
  • the resolution of the diluter 1 is in the range of 3000 to 24'000 steps per full stroke.
  • the tubing 5 has an internal diameter of 0.1 - 1.0 mm and the pipetting needle 4 has an internal diameter smaller than 0.6 mm at its free end and preferably smaller than 0.4 mm.
  • the pipetting needle 4 is coated externally with teflon, paraffin or fat or is silanized.
  • the coating produces a hydrophobic surface which results in a preciser break-off of the liquid from the tip of the needle 4.
  • the portion 12 of said tubing 5 located within said impulse generator 6 is made of a different material then the tubing 5.
  • the tubing 5 is generally made of polytetrafluorethylene (PTFE)
  • the portion 12 is made of silicone.
  • the application of the impulse indicated by hammer 20 on said portion 12 made of silicone gives optimum results.
  • Tubing 5 has an internal diameter of 0.1 - 1.0 mm.

Claims (20)

  1. Procédé pour le pipetage automatisé de volumes de liquide inférieurs à 10 µl au moyen d'un dispositif de pipetage pour l'aspiration et la distribution d'un liquide, caractérisé en ce que les étapes ci-dessous sont exécutées :
    A) une impulsion J d'au moins 0,1 N.s est appliquée au liquide contenu dans le dispositif de pipetage;
    B) après l'application de ladite impulsion, le liquide résiduel est avancé dans le parcours liquide;
    C) un volume défini de liquide, inférieur à 10 µl, est délivré par un dilueur (1) dans ledit parcours de liquide, ledit dilueur (1) ayant une sortie de liquide comprenant une seringue (2) et un clapet (3), ladite seringue (2) comprenant un piston (11) et un entraínement de piston (9);
    D) une impulsion supplémentaire J d'au moins 0,01 N.s est appliquée sur le liquide présent dans le dispositif de pipetage, suite à quoi
    E) ledit volume défini de liquide, inférieur à 10 µl, est distribué par le dispositif de pipetage.
  2. Procédé selon la revendication 1, caractérisé en ce que l'amplitude de ladite impulsion J appliquée sur le liquide dans le dispositif de pipetage est comprise entre 0,01 et 1,00 N.s.
  3. Dispositif de pipetage automatisé de volumes de liquide inférieurs à 10 µl, comprenant :
    A) un dilueur (1) présentant une sortie de liquide et comportant une seringue (2) et un clapet (3), ladite seringue (2) comportant un piston (11) et un entraínement de piston (9);
    B) une aiguille de pipetage (4); et
    C) un tube (5) reliant ladite sortie de liquide dudit dilueur (1) à ladite aiguille (4), ledit tube (5) et le dilueur (1) contenant un liquide qui peut être comprimé moins aisément que l'air,
       caractérisé en ce que le dispositif comporte en outre :
    D) un générateur d'impulsions (6) connecté dans ledit dispositif et couplé audit liquide présent dans ledit tube (5) pour générer et appliquer sur ledit liquide présent dans ledit tube (5) des impulsions J comprises dans la plage de 0,01 à 1,00 N.s, pour séparer le liquide de ladite aiguille (4); et
    E) ladite seringue (2) présente un volume compris dans la plage de 10 à 500 µl.
  4. Dispositif selon la revendication 3, caractérisé en ce que ledit générateur d'impulsions (6) est situé entre ledit clapet (3) dudit dilueur (1) et ladite aiguille (4).
  5. Dispositif selon l'une des revendications 3 à 4, caractérisé en ce que ledit générateur d'impulsions (6) est un générateur d'impulsions mécanique.
  6. Dispositif selon la revendication 5, caractérisé en ce que ledit générateur d'impulsions (6) comprend un noyau aimanté.
  7. Dispositif selon la revendication 5, caractérisé en ce que ledit générateur d'impulsions (6) comprend une vanne de passage à solénoïde.
  8. Dispositif selon l'une des revendications 3 à 4, caractérisé en ce que ledit générateur d'impulsions (6) est un générateur d'impulsions par induction.
  9. Dispositif selon la revendication 8, caractérisé en ce que ledit générateur d'impulsions (6) comporte un piézo-élément.
  10. Dispositif selon l'une des revendications 3 à 4, caractérisé en ce que ledit générateur d'impulsions (6) comporte une source d'ultrasons.
  11. Dispositif selon la revendication 3 à 4, caractérisé en ce que ledit générateur d'impulsions (6) comporte une source de chaleur.
  12. Dispositif selon l'une revendications 3 à 11, caractérisé en ce que ledit générateur d'impulsions (6) génère des impulsions J de 0,01 à 0,10 N.s.
  13. Dispositif selon l'une des revendications 3 à 12, caractérisé en ce que la seringue (2) présente un volume compris dans la plage de 100 à 400 µl.
  14. Dispositif selon la revendication 13, caractérisé en ce que la seringue (2) présente un volume compris dans la plage de 200 à 300 µl.
  15. Dispositif selon l'une des revendications 3 à 14, caractérisé en ce que la résolution du dilueur (1) est comprise dans la plage de 3 000 à 24 000 pas par course complète.
  16. Dispositif selon l'une des revendications 3 à 15, caractérisé en ce que ledit tube (5) est réalisé en polytétrafluoroéthylène (PTFE).
  17. Dispositif selon l'une des revendications 3 à 16, caractérisé en ce qu'une partie (12) dudit tube (5) située à l'intérieur dudit générateur d'impulsion (6) est réalisée en silicone.
  18. Dispositif selon l'une des revendications 3 à 17, caractérisé en ce que ledit tube (5) présente un diamètre intérieur de 0,1 à 1,0 mm.
  19. Dispositif selon l'une des revendications 3 à 18, caractérisé en ce que ladite aiguille de pipetage (4) présente à son extrémité libre un diamètre intérieur inférieur à 0,6 mm, et de préférence inférieur à 0,4 mm.
  20. Dispositif selon l'une des revendications 3 à 19, caractérisé en ce qu'à l'extérieur, ladite aiguille de pipetage (4) est dotée d'un revêtement hydrophobe, de préférence d'un revêtement en Téflon, en paraffine ou en graisse, ou d'un revêtement silanisé.
EP96937277A 1995-11-01 1996-10-29 Pipettage automatise de petits volumes Expired - Lifetime EP0876219B1 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US08/551,325 US5763278A (en) 1995-11-01 1995-11-01 Automated pipetting of small volumes
US551325 1995-11-01
PCT/EP1996/004691 WO1997016251A1 (fr) 1995-11-01 1996-10-29 Pipettage automatise de petits volumes

Publications (2)

Publication Number Publication Date
EP0876219A1 EP0876219A1 (fr) 1998-11-11
EP0876219B1 true EP0876219B1 (fr) 2000-05-31

Family

ID=24200804

Family Applications (1)

Application Number Title Priority Date Filing Date
EP96937277A Expired - Lifetime EP0876219B1 (fr) 1995-11-01 1996-10-29 Pipettage automatise de petits volumes

Country Status (7)

Country Link
US (1) US5763278A (fr)
EP (1) EP0876219B1 (fr)
AT (1) ATE193464T1 (fr)
DE (1) DE69608705T2 (fr)
ES (1) ES2150146T3 (fr)
PT (1) PT876219E (fr)
WO (1) WO1997016251A1 (fr)

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EP3450020A1 (fr) 2017-09-01 2019-03-06 Eppendorf AG Dispositif de microdosage permettant le dosage de plus petits échantillons de fluide
EP3485974A1 (fr) 2017-11-17 2019-05-22 Eppendorf AG Dispositif de microdosage permettant le dosage de plus petits échantillons de fluide

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WO2019043159A1 (fr) 2017-09-01 2019-03-07 Eppendorf Ag Dispositif de microdosage pour le dosage de micro-échantillons de fluide
EP3485974A1 (fr) 2017-11-17 2019-05-22 Eppendorf AG Dispositif de microdosage permettant le dosage de plus petits échantillons de fluide
WO2019096993A1 (fr) 2017-11-17 2019-05-23 Eppendorf Ag Dispositif de microdosage pour le dosage de micro-échantillons de fluide

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ES2150146T3 (es) 2000-11-16
EP0876219A1 (fr) 1998-11-11
PT876219E (pt) 2000-11-30
WO1997016251A1 (fr) 1997-05-09
DE69608705T2 (de) 2001-02-08
DE69608705D1 (de) 2000-07-06
US5763278A (en) 1998-06-09
ATE193464T1 (de) 2000-06-15

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